The role of alpha2delta in neuronal VGCC modulation

描述(由申请人提供)： 这项工作的目标是更好地了解辅助α2β亚基如何调节神经元电压门控钙通道(VGCC)，从而影响神经元迁移(Komuro，et。Al，1998)，突触发生(bahls，et.和神经传递(邓拉普，1995)。神经元VGCC由α1孔亚基和辅助亚基、α2β、β和可能的伽马亚基组成，它们以不同的方式调节通道特性(Catterall，2000)。异源表达系统的研究表明，α2β亚基与α1结合，并通过改变VGCC的膜转运和生物物理性质来调节通道功能(Klugbauer，et.Al，2003)。然而，Alpha2Delta在体内调节成孔Alpha1亚基的功能还不是很清楚。已分离到几个在果蝇α2Delta亚基中存在遗传损伤的直夹式(Stj)突变体。对stj突变体的初步电生理和EM研究提示囊泡融合存在缺陷。我建议研究果蝇α2Delta在体内的功能，重点是它在调节神经元VGCC和神经传递方面的作用。最初，我将生成Alpha2Delta的零等位基因，以帮助确定孤立突变的基因特征。然后，我将使用荧光标记的α1孔亚基调查VGCC在stj突变体中的贩运。此外，我将使用钙成像来确定Alpha2Delta是否调制通道功能。最后，我将探讨α2Delta和β之间的相互作用是否对于正常的VGCC功能是必要的。

DESCRIPTION (provided by applicant): The goal of this work is to better understand how auxiliary alpha2delta subunits modulate neuronal voltage-gated calcium channels (VGCCs) which affect neuronal migration (Komuro, et. al,1998), synaptogenesis (Bahls, et. al., 1998), and neurotransmission (Dunlap, 1995). Neuronal VGCCs consist of an alpha1 pore subunit and auxiliary subunits, alpha2delta, beta, and possibly gamma, which differentially regulate channel properties (Catterall, 2000). Studies in heterologous expression systems suggest that alpha2delta subunits associate with alpha1 and modulate channel function by altering the membrane trafficking and biophysical properties of VGCCs (Klugbauer, et. al, 2003). However, the in vivo function of alpha2delta in regulating pore-forming alpha1 subunits is less clear. Several straightjacket (stj) mutants with genetic lesions in the Drosophila alpha2delta subunit have been isolated. Initial electrophysiological and EM studies of stj mutants hint at defects in vesicle fusion. I propose to study the in vivo function of Drosophila alpha2delta, focusing on its role in regulating neuronal VGCCs and neurotransmission. Initially, I will generate null alleles of alpha2delta to aid in the genetic characterization of isolated mutations. I will then investigate VGCC trafficking in stj mutants using a fluorescent-tagged alpha1 pore subunit. In addition, I will determine whether alpha2delta modulates channel function using calcium imaging. Finally, I will explore whether interactions between alpha2delta and beta are necessary for proper VGCC function.